Apparatus and method for fluid collection and partitioning

ABSTRACT

Partitioning assemblies and partitioning or seal members, utilized with containers (adapted to serve as fluid specimen collection or fluid-retaining tubes) for effecting partitioning of two differing-density fluid phases of a centrifugally separated fluid specimen, at a position not lower than the fluid phase interface, wherein the partitioning members include a separating amount of a gel-like material. This gel-like material, by having a specific gravity intermediate those of the separated fluid phases, is adapted to move within the container in response to centrifugal force, only to the vicinity of the fluid phase interface. The gel-like material thereupon is further adapted to make a transversely continuous semi-rigid contact seal with an annular portion of the container inner surface to thereby effect a seal that partitions the fluid phases. The gel-like material may also be used in combination with a spool member having a container-contacting outer surface and a central axial orifice, with the gel-like material making a transversely-continuous contact seal within the spool central axial orifice. Three-phase partitioning may also be accomplished by using first and second gel-like materials having specific gravities intermediate those of the first-second and second-third differing-density phases, respectively. The partitioning or seal members may also be utilized in closed system (evacuated) fluid collection tubes or may be hand inserted into opened (atmospheric pressure) tubes after specimen collection. Also set forth is a method for effecting partitioning of centrifugally separated fluid phases within a container.

llnite States Patent Zine, ,lr.

[ Dec. 3, 19M

[ APPARATUS AND METHOD FOR FLUID COLLECTION AND PARTITIONING Anthony R.Zine, Jr., Corning, NY.

[73] Assignee: Corning Glass Works, Corning,

[22] Filed: Dec. 11, 1972 [21] Appl. No.: 314,270

[75] Inventor:

[52] US. (11...... 210/83, 2l0/DIG. 23, 2l0/DIG. 24,

233/1 A [51] Int. Cl B0101 21/26 [58] Field of Search 23/258.5; 106/287SB;

210/65, 83, 84, 512, DIG. 23;

Primary ExaminerCharles N. Hart Assistant Examiner-Robert G. MukaiAttorney, Agent, or FirmBurton R. Turner; Clarence R. Patty, Jr.

[57] ABSTRACT Partitioning assemblies and partitioning or sea] members,utilized with containers (adapted to serve as fluid specimen collectionor fluid-retaining tubes) for effecting partitioning of twodiffering-density fluid phases of a centrifugally separated fluidspecimen, at a position not lower than the fluid phase interface,wherein the partitioning members include a separating amount of agel-like material. This gel-like material, by having a specific gravityintermediate those of the separated fluid phases, is adapted to movewithin the container in response to centrifugal force, only to thevicinity of the fluid phase interface. The gel-like material thereuponis further adapted to make a trans' versely continuous semi-rigidcontact seal with an annular portion of the container inner surface tothereby effect a seal that partitions the fluid phases. The gellikematerial may also be used in combination with a spool member having acontainer-contacting outer surface and a central axial orifice, with thegel-like material making a transversely-continuous contact seal withinthe spool central axial orifice. Three-phase partitioning may also beaccomplished by using first and second gel-like materials havingspecific gravities intermediate those of the first-second andsecondthird differing-density phases, respectively. The partitioning 0rseal members may also be utilized in closed system (evacuated) fluidcollection tubes or may be hand inserted into opened (atmosphericpressure) tubes after specimen collection. Also set forth is a methodfor effecting partitioning of centrifugally separated fluid phaseswithin a container.

3 Claims, 10 Drawing Figures APPARATUS AND METHOD FOR FLUID COLLECTIONAND PARTITIONING BACKGROUND OF THE INVENTION 1. Field of the InventionThis invention relates to an apparatus and method for the collection andpartitioning of at least two phases of a muIti-phase fluid within acontainer. More specifically, it pertains to the collection of wholeblood and, after the separation thereof, the partitioning of blood serumor blood plasma from the blood cells. If desired further fractionatingand partitioning of, for example, the blood serum may be accomplished.

2. Prior Art In the standard evacuated blood sampling tubes, such as thesystem illustrated in US. Pat. No. 2,460,641 to Kleiner, a glass tubehas one permanently closed end and the other end is closed by a rubberstopper having a pair of opposite top and bottom axial recessesseparated by an intermediate diaphragm. A cup-like holder having adouble ended hollow needle, with one end terminating axially within theholder and the other end terminating axially outside the holder, is usedto receive the stoppered end of the glass tube, with the inner needleend being adapted to extend through the stopper diaphragm into theevacuated tube. The outer needle end is injected into the patients veinand then, by forward thrust on the tube, the puncturing of the stopperdiaphragm is completed to withdraw theblood. When the desired quantityof blood has been collected in the tube, the filled tube is removed fromthe cup-like holder thereby obtaining a stopper-sealed collection tubehousing a blood sample.

Blood or another fluid collected in the previouslydescribed collectiondevice is then generally taken to the laboratory for processing. Thecontents may be utilized as whole blood or separated into a lighterphase (serum or plasma) and a heavier phase (cells). If, for example, itis desired to obtain blood serum (after an initial time period duringwhich the filled tube assembly is allowed to stand) the filled tubeassembly is placed into a centrifuge which completes separation into twoblood phases. Disposed at the bottom of the tube will be a heavy phaseor high density portion of the fluid consisting of packed red bloodcells, while disposed at the upper part of the tube will be the lighterphase or low density portion of the fluid which is blood serum. Theseparated serum is then analyzed, generally after first being removedfrom the tube assembly by decanting and/or siphoning.

It is well known that once the blood phases are separated, if thelighter phase is not removed from the tube within a short time,interaction will occur between the separated phases and inaccurate testresults will be obtained. In addition, even if the lighter phase ispresently removed from the container there are the hazards ofcontamination of the sample and of possible mismarking of the removedsample. Furthermore, there are also hazards to the laboratory personnelwho may be ex posed to disease-carrying blood samples containing, forexample, hepatitic serums.

Coleman, in US. Pat. No. 3,508,653, made an advance over the bloodsampling tube of Kleiner by introducing and attaching a resilient pistondirectly beneath the tube closure or stopper, with the piston beingadapted to be punctured during the initial filling of the sampling tube.After initial centrifugation, in order to obtain the desired blood phaseseparation, and in re sponse to further centrifugal force, the piston isdesigned to move downwardly through the light blood phase, with thepiston being adapted to permit upward flow of the light phasetherearound, i.e., between the container inner wall surface and theouter peripheral surface of the piston. The piston, which has a wiperportion that makes an initial sealing contact with the container innersurface, loses this sealing contact during its downward movement (topermit the flow of fluid therearound) and thereafter is designed to makea final sealing contact with the container inner surface at a positionnot lower than a position intermediate the separated phases by stoppingthe downward movement by terminating the applied force. In addition, thepiston, which is initially detachably secured to the stopper, requirespassageway means and a vent opening therewithin to facilitate thepassage of gases to permit descent of the piston but resist the passageof fluids therethrough.

While the Coleman device provides a unitary sealing member between theblood cells and the plasma or serum, it does have several shortcomings.The piston and stopper must be held in intimate contact with each other,otherwise blood which flows into any space between them during the tubefilling operation will remain above the piston, and the blood cells willcontaminate the lighter phase. Once these blood cells find their wayabove the piston wiper, they cannot be separated, since no mechanism ormethod has been provided to permit them to move below the piston.

In addition, there are no positive means incorporated into the Colemandevice to prevent blood cells from moving upward past the piston wiper.Actual observations in the laboratory confirm that in spite of thegeneral downward movement of the heavy phase, due to the influence ofcentrifugal force, some blood cells do indeed become caught up in thefast'moving light phase stream and are carried past the piston wiperinto the upper chamber of the tube. As noted, once the cells find theirway above the piston wiper, there is no way to return them to the lowerportion of the tube.

Since the introduction of the blood sample into the tube may also permitsome airto enter the tube upon withdrawal from the patient and sincesome gases are evolved from the blood sample, they must be vented frombelow the piston to eliminate the retarding effect they will have on thedownwardly moving piston through a buoyancy effect. While Coleman speaksof incorporating a vent opening into the piston design, actualexperience has shown that the vent cannnot readily be incorporated intothe design at manufacture but is preferably made by the technicianduring the blood drawing operation, thereby putting the burden ofcreating a satisfactory vent upon the skill of the operator. The needlepuncture in the piston diaphragm (for the filling of the tube) serves asa vent for air and gases during piston descent. An improperly punctureddiaphragm vent may either refuse to operate at all or may rupture andblow out when the piston impacts the fluid surface during centrifugationand thus completely loses its ability to act as a seal between the lightand heavy blood phases during piston descent. In either instance,unfortunately the separation step becomes aborted.

Lawhead, in US. Pat. application Ser. No. 228,573 filed Feb. 23, 1972,made an advance over the method and apparatus of Coleman by introducingspools or partitioning assemblies for use with rigid tubular containersfor effecting either the physical or complete physical and chemicalpartitioning of two centrifugally separated fluid phases. These spoolshave a central axial orifice, a resilient, annular, container-contactingwiper portion and an integral annular skirt portion. By having specificgravities intermediate those of the separated fluid phases, the spoolsare adapted to move downwardly in the tubular containers, in response tocentrifugal force, only to the vicinity of the fluid phase interface,with fluid flow occurring freely only through the spool central axialorifice. Partitioning of the separated phases is effected by thecombination of the spool in conjunction with either a natural plug ofthe heavy phase fluid or afloat me'mber'having a similar specificgravity.

While the Lawhead device produces excellent sealing between theseparated phases, it does require different diameter parts for differentdiameter tubes, which of course is an economic disadvantage in a lowunit cost system.

Weichselbaum, in US. Pat. No. 3,464,890, sets forth a method ofseparating plasma from whole blood which comprises bringing into contactwith the blood a separating amount of inert particulate material, e.g.,polystyrene beads having a coating of anti-coagulant and having aspecific gravity intermediate that of plasma and blood. This loosematerial is placed into the blood sample prior to phase separation andupon separation these particles tend to establish a barrier between theplasma and cells. This system, however, will not tolerate any subsequentjarring or unusual motion since this will tend to destroy the barrier.Furthermore, this system will not tolerate shipping and cannot beutilized for mailing to testing laboratories.

Adler, in US. Pat. No. 3,647,070, sets forth a method and apparatus fora barrier at the interface between plasma and packed cells incentrifuged blood samples, which barrier means are adapted to sinkthrough the plasma layer, and upon being wetted and expanded by theplasma, expanded into firm contact with each other and the walls of thecontainer to form a barrier. While this system appears to be quiteworkable, it is limited to post centrifugation insertion of the barriermeans which is a definite disadvantage from the cost, time andcontamination standpoint. I

The use of silicones for centrifuge fractionating of blood samples iswell known and is set forth in articles by Seal, S. H. in Cancer. 195912:590-595; McCrea, L. E. in J. ufUrol. 1961. 85: 1006-1010; as well asMorgan M. C. and Szafir j. J. in Blood. 1961. 18:89-94. These articlesbasically describe the use of silicone fluids (blended to specificgravities intermediate those of the two phases sought to be separated)with blood samples, with the silicone fluid, upon centrifugation,forming a fluid barrier between the desired two phases. However, sincethe barrier is only a fluid barrier the desired phase cannot be removedby decanting and even in pipetting there is a problem of possiblecontamination of the removed phase with silicones. Furthermore, theseliquid barriers will neither tolerate any subsequent jarring nor arethey adaptable to shipping. 1

SUMMARY OF THE INVENTION The instant invention, both in terms ofapparatus and method, responds to each of the previously-described priorart shortcomings in a manner so as to completely eliminate any furtherconcern regarding such problems.

The several embodiments of the partitioning assemblies and partitioningor seal members of this invention are utilized with containers that areadapted to serve as fluid collection or fluid retaining tubes.

The partitioning or seal members include a predetermined or separatingamount of a gel-like material, preferably hydrophobic, substantiallythixotropic and generally inert to the separated fluid phases that areto be partitioned. This gellike material, such as a mixture of asilicone fluid and hydrophobic silicon dioxide powder, which has aspecific gravity intermediate those of the fluid phases, is positionedwithin the container either before or after fluid collection. Due to itsspecific gravity, the gel-like material is adapted to move within thecontainer in response to centrifugation, with the gel-like materialbeing adapted to stop moving when it reaches the vicinity of the fluidphase interface. The gel-like material thereupon is further adapted tomake a transversely-continuous, semi-rigid, contact seal with an annularportion of the container inner surface, thereby effecting a seal thatphysically and chemically partitions the fluid phases.

While the gel-like material may be used by itself to form a semi-rigidpartitioning or seal member, it may also be used in combination with aspool member having a container-contacting outer surface portion and acentral axial orifice. The spool member, which is preferably initiallypositioned below the container stopper or closure, by having a specificgravity that is intermediate those of the separated fluid phases, isadapted to move downwardly within the container in response tocentrifugal force. The fluid phases flow freely only through the spoolcentral axial orifice, with the spool being adapted to stop movingdownwardly when it reaches the vicinity of the fluid phase interface.The gel-like material, which in this combination is preferably initiallylocated adjacent to the bottom of the container, by reason of itsspecific gravity, moves upwardly within the container and is adapted tomake a transversely continuous semi-rigid contact seal with at least anannular surface portion of the spool central axial orifice.

The partitioning or seal members of this invention may also be utilizedto partition at least three differing density phases of a separatedmulti-phase fluid specimen at positions substantially at the interfacesof these fluid phases. This three-phase partitioning may be accomplishedby using first and second gel-like material having specific gravitiesintermediate those of the firstsecond and second-third differing densityphases, respectively. These gel-like materials are adapted to makeseparate transversely-continuous, semirigid, contact seals withdifferent annular portions of the container inner surface therebyeffecting seals that partition the three separated phases.

.The partitioning assemblies and partitioning or seal members of thisinvention may be utilized in several different operational sequences.One operational sequence applies specifically to a fluid collection andpartitioning assembly that is intended to remain closed (vacuum sealed)from the time of manufacture through sampling, preparation andcentrifugation of its contents until the lighter phase is removed aftercentrifugation.

In another operational sequence, the partitioning or seal member ishand-inserted or dispensed into an opened collection tube (i.e., atatmospheric pressure) after sample collection, prior to centrifugation.

In the closed system concept sequence, the gel-like material may bepositioned anywhere within the collection tube, while in thehand-insertion concept sequence, the gel-like material is preferablydispensed into the tube either as a floating capsule or positioned onthe side of the tube below the tube closure.

When three or more phase partitionings are desired, both closed systemand hand insertion concept sequences, as well as combinations thereof,may be employed, with one or more centrifugation steps being required.

One method of establishing the partitioning of heavier phase from thelighter phase of a centrifugally separated fluid specimen within acontainer involves providing the container with a predetermined amountof a gel-like material having a specific gravity intermediate those ofthe separated phases. Moving the gel-like material within the containerthrough at least one of the fluid phases (in response to centrifugalforce) establishes a flow of at least one of the fluid phases within thecontainer. A transversely-continuous semi-rigid contact seal isestablished with an annular portion of the container inner surface whenthe gellike material reaches a position in the vicinity of the fluidphase interface thereby partitioning the lighter and heavier fluidphases. Thereafter the applied force is terminated.

Other advantages and features of the instant invention will beunderstood from the following description in conjunction with theattached drawings.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. I illustrates one of the fluidcollection and parti tioning assemblies of this invention, ready foruse, with the partitioning or seal member in the form of a gel-likematerial being initially position adjacent to the normally closed end ofthe tubular container.

FIG. 2 is the assembly of FIG. I after theintroduction of a homogenizedfluid sample thereinto.

FIG. 3 illustrates the assembly of FIG. 2 shortly after the start ofcentrifugation, which begins to separate the homogenized sample into atleast two differing-density fluid phases, with the gel-like materialbeginning to move away from its initial position.

FIG. 4 illustrates that in the assembly of FIG. 3, as centrifugationcontinues, the gel-like material is approaching the interface betweenthe two differingdensity fluid phases.

FIG. 5 illustrates the assembly of FIG. 4i upon the completion ofcentrifugation, with the gel-like material being located at theinterface between the differingdensity fluid phases and making atransversely continuous contact partition or seal to thereby physicallyand chemically partition the two separated phases.

FIG. 6 illustrates another embodiment of the fluid collection andpartitioning assemblies of this invention, having a spool poised beneaththe closure member of the container and having a predetermined amount ofgel-like material positioned adjacent to the naturally closed end of thecontainer, with the differing density fluid being disposed therebetween.

FIG. 7 illustrates the assembly of FIG. 6 upon the completion ofcentrifugation, with the spool and gellike material being located at theinterface between the differing density fluid phases and coacting tomake a transversely-continuous contact seal to thereby partition thesephases.

FIG. 8 is a sectional view, partially broken away. of one of the fluidcollection and partitioning assemblies of this invention wherein thegel-like material is dispensed into the fluid collection assembly afterthe fluid collection is completed.

FIG. 9 is a sectional view, partially broken away, of another embodimentof the fluid collection and partitioning assemblies of this invention,having separatelypositioned first and second gel-like materials ofdiffering densities and at least a three-phase fluid specimen disposedtherebetween.

FIG. It) illustrates an assembly, such as that of FIG. 9, upon thecompletion of centrifugation, with the first and second gel-likematerials being located in the form of transversely continuouspartitioning members or contact seals at the interfaces between thefirst-second and the second-third differing density phases,respectively.

' DESCRIPTION OF THE PREFERRED EMBODIMENTS Referring now to the drawings:in detail, FIGS. 1-5 illustrate one of the fluid collection andpartitioning assemblies or container assemblies of this invention bothin terms of the various components in correct relation ship to eachother as well as the operational sequence of the various parts thereof.

FIGS. I-5 depict a fluid collection and partitioning assembly, morespecifically, a blood collection and partitioning assembly or containerassembly II consisting of a container or collection tube 12; apredetermined amount of a gel-like material 30; and a stopper or closure2Q; all of which will now be described in more detail.

Collection tube I2, which is preferably made of glass, plastic or othermaterial, and which is preferably also transparent, has a normallyclosed. bottom end M and an open upper end I6 for receiving aself-sealing stop per or closure 20 formed of medical grade butyl rubberor other suitable material. Closure 20 may be of the shape and materialdescribed herein or it may be of other suitable known types. Stopper 20as shown, is shaped so as to have a flanged end. 22 which abuts andoverlies annular end face I8 of collection tube open end 16. Stopper 20is further provided with a diaphragm or septum 24lwhich forms atransverselycontinuous seal with an annular surface portion of tubeinner wall surface I3. Stopper 20, together with collection tube I2,defines a sealed, closed fluid receiving chamber 26, which in thearrangement shown in F I6. I is adapted, (after previously having beenevacuated) to maintain a negativepressure (vacuum) of about 24 inches Hgfor an extended period of time. Thus. stopper 26), serves as a sealingclosure to preserve the interior vacuum and provides a septum 24 throughwhich the sampling needle (not shown) can reach chamber 26 withoutdestroying its integrity. No invention is claimed for either thepreviously described collection tube 12 or stopper 20, per se.

Again, as shown in FIG. I, a predetermined amount of a gel-like material30 preferably is initially positioned adjacent to closed end M of tube112. This pre determined or separating amount (such as about 1 ml) ofgel-like material 30 preferably is hydrophobic, thixotropic andgenerally inert to body fluids. One example of such a gel-like materialis a mixture ofa silicone fluid and very fine hydrophobic silicondioxide powder. I-Iydrophobic silicon dioxide (SiO may be defined assilicon dioxide that is treated so as to repel water, with one exampleof a hydrophobic silicon dioxide powder being Silanox 101 (manufacturedby the Cabot Corporation of Boston, Massachusetts and described in Cabotbrochure SGEN-l) hydrophobic fumed silicon dioxide, which is a fumedsilicon dioxide having trimethyls'ilyl groups bonded to the surfacethereof. Another example of a hydrophobic silicon dioxide powder isAEROSIL R972 (sold by DEGUSSA INC. Pigments Div., New York, NY. anddescribed in Technical Bulletin 31 wherein the silicon dioxide isrendered hydrophobic by reacting the silanol groups on the surface withdimethyl dichlorsilane.

Silicon fluid may be defined as a polysiloxane liquid such as forexample DOW Corning 360 Medical Fluid (a dimethyl polysiloxane liquidmanufactured by the DOW CORNING Corporation of Midland, Michigan anddescribed in DOW CORNING Bulletins CPO-1072, March, 1972 and CPO-158-1,March 1972). Other examples of silicone fluids are DOW CORNING 200 and510 (a methylphenyl polysiloxane) fluids.

The following specific example of a gel-like material is given inillustration of the present invention and is not intended to be limitingon the invention. One hundred grams of DOW CORNING 360 Medical Fluid(350 centistokes; specific gravity about 0.97) were mixed with 15.7grams of Silanox 101 silanemodified silicon dioxide (specific gravityabout 2.2) to produce 115.7 grams of a gel-like material having aspecific gravity of about 1.05.

Table 1 illustrates, among others, a number of mixtures of gel-likematerials that may be utilized in this invention:

Sample about 1.06, with the preferred range being 1.04-1.055.

With reference to one of the operational sequences of this invention,FIG. 1 illustrates fluid collection and partitioning assembly 11, readyfor use, with stopper 20 together with collection tube 12 defining asealed, closed evacuated fluid receiving chamber 26. Con tained withinchamber 26 is gel-like material 30 which is positioned adjacent tonormally closed end 14 of tube 12.

The FIG. 2 assembly depicts the FIG. 1 assembly with the addition of amulti-phase fluid sample 34, such as whole blood. After a correctvenipuncture has been made on the patient, the inner or butt end of theneedle (not shown) is pushed through stopper diaphragm portion 24,thereby permitting the vacuum within the assembly to draw blood freelyinto tube 12.

FIG. 3 illustrates the assembly of FIG. 2 shortly after the start ofcentrifugation which begins to separate homogeneous fluid sample 34 intoa lighter phase 38 and a heavier phase 42. The interface 44 betweenlighter and heavier phases 38 and 42, respectively, is shown, for thesake of clarity, in the form of a dash on either 'side of tube 12.During centrifugation, heavier phase tial position.

The FIG. 4 assembly shows the FIG. 3 assembly, as centrifugationcontinues, with gel-like material 30 still in an elongated form, but nowfully removed from its initialposition, with the upper end of gel-likematerial 5111- Viscosity Grams of Grams of Grams of Resulting No. cone(centistokes) Silicone S-101 R-972 S.G.

Table l DC-360 is DOW CORNING 360 Medical Fluid. DC-SlO is DOW CORNING510 Fluid. S-lOl is Silanox" 101 hydrophobic 510,. R-972 is DEGUSSA R972 hydrophobic Si0,.

It should be noted that the specific gravity (S.G.) of whole blood is1.05l.06 while the S.G. of the light phase (blood serum) is 1.02-1.03and the S.G. of the heavy phase (blood cells) is 1.08-1.09. Thereforethe specific gravity of gel-like material 30 has to be below that of theheavy phase and above that of the light phase, i.e., generally in therange from about 1.035 to 30 being located in the vicinity of fluidphase interface 44. It should be noted that a thin layer 32 of gel-likematerial remains at its initial position, i.e., at tube bottom 14.

It should be noted that due to the resilience of the gel-like material30, movement of fluid can occur in either direction. i.e., red bloodcells. fibrin or other heavy-phase bodies can usually move downwardlythrough gel-like material 30 under the persuasion of centrifugal force.At the same time, any lighter phase fluid remaining below gel-likematerial 30, again under the persuasion of centrifugal force, canusually move upwardly through material 30. The operation of the instantinvention is such that it does not differentiate be tween gases andliquids and permits both to flow through material 30 without prejudice.The flow, either of gases or liquids, is neither restricted norotherwise influenced in any way by the gel-like material 30. Each phaseis free to seek its own flow path and its ultimate position within tubeI2 is influenced solely by the persuasion of centrifugal force.

FIG. illustrates the assembly of FIG. 4- upon the completion ofcentrifugation, i.e., all the parts are now in final position. Upon thecompletion of centrifugation the maverick lighter components or cells ofheavier phase 42 (previously in or above material 34)), still having aspecific gravity greater than that of material 30, have now eased intoor through material 30., with mate rial resting at a density levelequivalent to its own specific gravity. As shown in FIG. 5 member 3t)has now consolidated so as to make a transverselycon'tinuous semi-rigidcontact seal or partitioning member 48 with an annular surface portionof tube inner surface 13. If the homogenized test fluid 34 is wholeblood, then the heavier phase 42 is now blood cells and the lighterphase may be either blood serum or blood plasma, depending upon whetheror not the whole blood sample was coagulated or not coagulated,respectively.

It should be understood that the thickness or axial dimension of thetransversely-continuous contact seal made by partitioning or seal member4% is, among other things, of course also dependent upon the amount ofgel-like material that is initially introduced into tube 12. Inaddition, the seal need not be of uniform shape or thickness across itstransverse dimension 1 as long as it has at least onetransversely-continuous portion. Uniformity of the seal is influenced bysuch factors as the viscosity of the gel-like material, the amount ofmaterial present, the speed and type (horizontal or angleheadcentrifuge) of centrifugation (and resulting g-force) as well as thecentrifugation time.

It should be noted that gel-like material 30, which makes uptransversely-continuous semi-rigid seal member 48, is substantiallythixotropic, i.e., at rest it acts substantially like a material in athixotropic state. It is not intended that this definition of thegel-like material, which also may be described as semisolid, semi-rigid,substantially non-flowable, or resistant to flow at rest, be alimitation on the invention herein described, since the behavior of thematerial is, at this time, not yet completely subject to a full exactingexplanation. It should suffice to say that gel-like material 30 appearsto have a very high viscosity, is thermoplastic in nature, will actsubstantially as a fluid during centrifugation and will again set up toa gel when allowed to stand.

In the form of seal-member 48, gel-like material 30 is substantiallyrigid and allows decanting of the lighter fluid phase from the tube orcontainer 32 without disrupting its seal with the tube inner surface. Inaddition, the partitioned sample will readily tolerate subsequentjarring and is entirely adaptable to shipping (such as to a remotelaboratory for example).

While the previously-described examples of gellike materials 30 aremixtures of silicone fluids and silicone dioxide powders it must beunderstood that these mixtures are not to be considered as limiting thisinvention. Any gel-like material is useful in the context of thisinvention if it meets the following basic requirements:

1. Specific gravity (or density) intermediate between those of the twofluid phases sought to be sepa rated.

2. Non-interaction with the fluid phases sought to be separated.

3. Substantially non-flowable (semi-rigid) at rest.

In the previously-described examples. the silicon fluid may be thoughtof as a liquid or base material (an oil) and the silicon dioxide powderas a solid (a filler), with the latter serving both to adjust thespecific gravity of the former to the desired value and to gel the oil,i.e., to convert it into a semi-rigid gel-like material or grease (withthe terms gel-like and grease being used synonymously). Thus, as long asthey meet the previously noted three basic requirements, almost anyliquid and filler combination may be utilized, with examples of oilsincluding esters of polyacids (such as dioctylsebacate, dibutylphthalateand tributylphosphate) and min eral oils (hydrocarbons). Examples offillers include titania, zirconia, asbestos, wood flour and finelydivided organic polymers (such as polyethylene, polypropylene,fluorocarbons and polyesters, etc.) In addition, depending on thespecific gravity of the base material. the fillers may be used to eitherincrease or decrease the specific gravity of the former. Furthermore,again as long as the three basic requirements are met, the gellikematerial may be made up of but a single component (such as a silicone)material or may be mixtures of one or more base materials and one ormore fillers.

Up to this point the only operational sequence described has been onewherein gel-like material 30 is initially positioned adjacent to closedend 14 of tube 12, as shown in FIGS. 15. However, other initialplacements of material 30 are entirely possible, i.e., material 30 maybe placed anywhere within fluid receiving chamber 26. For example, asshown in FIG. 9, a predetermined amount of gellike material 30a may bepositioned on a portion of tube inner'surface 13 below stopper 20. Whenused in the sequence shown in FIGS. ll-S, in lieu of material 30, uponcentrifugation, gel-like material 3% (which is substantially similar tomaterial 30), by virtue of its specific gravity (l.035l.()6) will movedownwardly through lighter phase 38 (specific gravity 1024.03) andeventually rest at the density level equivalent to its own specificgravity. The end result, as shown in FIG. 5, will be substantially thesame regardless of whether material 30 moves up from tube bottom M ormaterial 31M moves down from the vicinity of stopper 20.

The operational sequences described up to now have been limited to afluid collection and partitioning assembly Ill consisting of collectiontube 12, stopper 2d and contact seal or partitioning member 48, whereintube 12 together with stopper 20 defines a sealed, closed, evacuatedfluid receiving chamber 26. This operational sequence, as shown in FIGS.1-5, applies specifically to a fluid collection and partitioningassembly that is intended to remain closed from the time of manufacture,through sampling, preparation and centrifugation of its contents untilthe lighter phase is to be removed after centrifugation. Of necessity,the gel-like material must be placed into the tube (prior to theevacuation thereof) at the factory. This sequence will hereinafter bereferred to as the closed system concept to differentiate it from a handor user insertion concept.

In an operational sequence utilizing the hand insertion concept, apredetermined amount of gel-like material such as for example 30a inFIG. 9 or 30b (also substantially similar to material 30) in FIG. 8 isdispensed into an opened collection tube after sample collection,preferably either after coagulation has been completed or after partialphase separation has been effected (upon completion of coagulation). Thegellike material can be inserted into an opened collection tube evenbefore coagulation has been completed, however, since blood cellsexhibit a tendency to harden on the walls of the opened tube it ispreferable to delay the opening of the collection tube until coagulationhas been completed therein.

With reference to the operational sequence utilizing the hand-insertionconcept, FIGS. 2 and 3, sans material 30, may be utilized to illustratea well-known evacuated blood collection tube assembly comprised ofcollection tube 12 and stopper 20. Once blood sample 34 has beenintroduced into this assembly and preferably eitherafter coagulation(FIG. 2) or after partial phase separation (FIG. 3), stopper 20 isremoved and gel-like material 30a (FIG. 9), or 30b (FIG. 8) is dispensedinto tube 12.' Thereafter, stopper 20, in accordance with good medicalpractice, preferably is placed back on tube 12 and centrifugation canbegin (FIG. 2, sans material 30) or be continued (FIG. 3, sans material30). Hereinafter, the operational sequence proceedsin a manner and witha result identical to that already described with reference to theclosed system concept. Substantially similar results are obtainedregardless of whether the gel-like material is dispensed directly intothe fluid sample, as is material 30b in FIG.

8, or positioned on an inner surface portion of the tube, as is material300 in FIG. 9.

FIGS. 6 and 7 disclose another embodiment of the fluid collection andpartitioning assemblies of this invention wherein a predetermined amountof gel-like material 30 coacts with a spool 52to effect completephysical and chemical partitioning of two differingdensity fluid phases.The assembly shown in FIG. 6, i.e., tube 12, stopper 22, gel-likematerial 30, fluid 34 and spool 52, can be the result of at least twodifferent concept sequences, namely: l a closed system concept whereinspool 52 and gel-like material 30, are both located in a sealed, closed,fluid receiving chamber 26 as shown in FIG. 1., into which fluid sample34 has thereafter been introduced, or (2) a hand-insertion conceptwherein spool 52 is introduced into a collection tube 12 (upon stopperremoval) after fluid sample 34 has been collected (as shown in FIG. 2).

Spool 52, which has an annular, generally cylindrically-shaped main bodyportion 54 having a diameter less than the inside diameter of collectiontube 12, also has an upper. outwardly-tapering, annular. resilient,wiper or outer surface portion 56 having a maximum outer free diametergreater than that of portion 54, with portion 56 being adapted tosealingly contact tube inner wall surface 13. Spool 54 also has a lowerskirt portion 58 and a central axial orifice 62. Spool 52 may be of thetype disclosed in co-pending US. Pat. application Ser.

No. 228,573 filed Feb. 23, 1972 (which is a continuation in part ofapplication Ser. No. 178,274 filed Sept. 7, 1971) and is also assignedto the assignee of this invention.

With reference to the operational sequence of the FIG. 6 and 7embodiments. FIG. 6 shows the fluid collection and partitioning assemblyimmediately prior to centrifugation, while FIG. 7 depicts the assemblyupon the completion of centrifugation. During centrifugation, gel-likematerial 30 behaves in the manner already described with reference toFIGS. 3 and 4 except that material 30 coacts with spool 52 to make atransverse ly-continuous contact seal to separate phases 38 and 42.Spool 52, which is preferably made .of a resilientmaterial such asmedical grade rubber, preferably has a specific gravity intermediatethose of the fluid phases to be separated (in the case of human bloodthe intermediate S.G.=l.035l.06). At the start of centrifugation, spool52, because of its specific gravity, starts to move downward, away fromthe vicinity of stopper 20, toward lighter phase 38, which in turn flowsupwardly through spool central axial orifice 62. It should be noted thatall fluid flow takes place through orifice 62 and no fluid is permitted,nor can it possibly take place, between the outer surface of spool 52and tube inner surface 13. Furthermore, fluid flow can occur throughorifice 62 in either direction, depending upon the initial position ofspool 52 relative to the various density components of the fluid whichare to be separated. The operation of spool 52 is such that it does notdifferentiate between gases or liquids and each phase is free to seekits own flow path and its ultimate position within tube 12 is influencedsolely by the persuasion of centrifugal force. Upon the completion ofcentrifugation (FIG. 7) the skirt portion 58 of spool 52 has enteredheavier phase 42 and gel-like material 30, again as a result of theapplied centrifugal force, has started to enter lighter phase 38 byextending at least partially through spool central axial orifice 62.Gel-like material 30 is adapted to make a transversely-continuouscontact seal member 64 with at least an annular portion of orifice 62.Thus spool 52 together with gel-like material 30 forms atransversely-continuous partitioning assembly 66 with an annular surfaceportion of tube inner surface 13. Basically, spool 52 acts as aconstriction within tube 12 since sealing of the differing densityfluids from one another at tube inner surface 13 has been continuous (byreason of spool wiper 56) since spool 52 began its descent through thefluid and the separated fluid phases have never been in contact witheach other in this area. Final sealing is accomplished within spoolcentral axial orifice 62 due to the action of gel-like material 30, andis purposefully designed to occur at or just above the fluid phaseinterface 44 to ensure the absence of any heavy phase components withinthe lighter phase sample. The exact positioning of gel-like material 30with reference to spool skirt portion 58 and orifice 62 depends upon theamount and viscosity of material 30 as well as the centrifugal forceapplied.

Up to this point the embodiments described have been limited to thepartitioning of two differing-density fluid phases of a centrifugallyseparated fluid specimen at the interface of the fluid phases. Often itmay be desirable to partition at least three differing-density phases ofcentrifugally separated muIti-phase fluid specimen at positionssubstantially at the interfaces of the differing-density fluid phases.This goal can be accom- 13% plished by utilizing n-l differing-densitygel-like materials to partition n number of differing-density fluidphases.

FIG. 9, which is a partially broken away sectional view of anotherembodiment of the fluid collection and partitioning assemblies of thisinvention, shows a first gel-like material 39, having a first specificgravity, adjacent to tube bottom I4 and a second gel-like material 30a,having a second specific gravity, attached to tube inner wall 13 in anarea below stopper 29. Art at least three-phase fluid 68 having first orheaviest density 70, second or intermediate density 72, and third orlightest density 74 fluid components is also contained within tube 12.Gel-like material 20 has a specific gravity intermediate those of firstand second fluid phase components 7t), 72, respectively, while material390 has a specific gravity intermediate those of second and third fluidphase components 72, 74 respectively.

FIG. 10 which illustrates an assembly, such as that of FIG. 9, upon thecompletion of centrifugation, with first and second gel-like materials39, 39a, being located in the form of transversely-continuous semi-solidpartitioning members or contact seals 48, lfla, between first-second(7tlt-72) and second-third (72-74) differing density fluid phases,respectively. As already previously described, gel-like material 30moves upward away from tube bottom 14 and material 30a moves downwardaway from the area below stopper 20, under the influence of centrifugalforce until they reach the fluid gradient levels, i.e., the interfacesclosest to their own specific gravity.

The end result, i.e., the partitioning of fluid phases 70, 72 and 74shown in FIG. 10 may be achieved in a number of different operationalsequences. In the closed system concept technique, both materials 30 and30a are contained (at tube bottom M and in the area below stopper 20,respectively) within a closed, evacuated fluid receiving chamber, intowhich fluid sample 68 is thereafter introduced (see FIG. 9). Asubsequent single centrifugation step produces the partitioning shown inFIG. Ill), with both materials 30 and 30a leaving thin layers ofgel-like materials 32 and 32a respectively at their initial positioningareas.

In the hand insertion concept technique, after fluid sample 68 has beencollected, a first gel-like material, having a first specific gravityintermediate those of heaviest and intermediate fluid phases 70, 72,respectively, is dispensed into collection tube 12, either in the shapeof material 30b (FIG. 8) or material 3th: (FIG. 9). Thereafter, theassembly is centrifuged a first time and this first gel-like materialforms transverselycontinuous partitioning or seal member 4% betweenphases 70 and 72. Then, a second gel-like material, having a secondspecific gravity intermediate those of the intermediate and lightestfluid phases, 72 and 74, respectively, is dispensed into collection tube112, again either in the shape of material 30b (FIG. 9) or material 30a(FIG. 9). After a second centrifugation, the second gel-like materialforms transversely-continuous partitioning or seal member Ma betweenphases 72 and 74.

Several combination closed system and hand insertion" concept techniquesare also possible. In one such combination, a first gel-like material,having a specific gravity intermediate those of heaviest and in--termediate phases 7t), 72, respectively, is contained within a closed,evacuated fluid receiving chamber (FIG. I). After the introduction offluid 68 a second gel-like material, having a specific gravityintermediate those of the intermediate and lightest phases 72, 74. re

spectively, is dispensed into collection tube 12 (either into collectiontube 12 (in either of the shapes as previously noted) and a secondcentrifugation step then forms a second transversely-continuous member48a between phases 72 and '74.

It should be noted that with any multi-phase fluid, having three or morephases, an initial separating of the heaviest phase from the remainingphases can be accomplished by means of either of the two phaseseparation techniques herein discussed, e.g. by means of partitioning orseal member 4 8 (FIG. 5) or partitioning assembly 66 (FIG. 7), usingeither the closed system or hand-insertion concept techniques. Furtherseparations of the remaining phases can thereafter be accomplished bysuccessively dispensing in gel-like materials of decreasing specificgravities and successively centrifuging the collection tube assembly.For example, a whole blood sample may be initially be separated intoblood cells and blood serum and thereafter, while remaining in thesamecontainer, the blood serum may be further fractionated into separatecomponents. Whole human blood has a given, generally quite uniform,specific gravity between 1.05 and 1.06. Centrifuged blood however hasmany layers or constituents of varying specific gravities, from theheaviest at the bottom to the lightest at the top, with the greatestvisible demarcation occurring at the serum/red cell interface.

The principles of this invention may be utilized in partitioningassemblies for fluids other than human blood, e.g., any fluid separableinto at least two differing density phases may be separated using agel-like material (or a gel-like material and spool combination) havinga specific gravity intermediate those of the phases sought to beseparated. Any gel or gel-like material that is hydrophobic,substantially thixotropic, and generally inert to the fluids to beseparated, may be utilized.

While the invention has been described in connection with possible formsor embodiments thereof, it is to be understood that the presentdisclosure is illustrative rather than restrictive and that furtherchanges or modifications may be resorted to without departing from thespirit of invention or scope of the claims which follow.

I claim:

11. A fluid collection and partitioning assembly for collecting aspecimen of blood within a sealed fluid collection chamber,centrifically separating the heavier and lighter fluid phases of saidblood specimen, and physically and chemically partitioning the separatedphases, comprising:

a. a container having an open end and a closed end;

b. gel-like means initially positioned within said container adjacentsaid closed end for forming a transversely continuous contact seal withan annular surface portion within said container at a subsee. subjectingsaid specimen and gel-like means to a quently formed interface betweensaid heavier and lighter phases;

said container and for defining a closed fluid collection chambercontaining said gel-like means therewithin, said closure means beingpierceable closure means for vacuum-sealing said open end of centrifugalforce to separate said fluid specimen into a heavier phase and a lighterphase and simultaneously move said gel-like means toward the interfaceof said phases; and

f. establishing a continuous semi-rigid gel-like seal across theinterior of said container between said by a needle for supplying bloodto said closed fluid collection chamber which is adapted to draw the 0blood specimen therewithin;

d. said gel-like means being a thixotropic material and including amixture of a fluid which is generally inert to body fluids and apowdered inorganic filler; and

heavier phase and said lighter phase within said container.

3. A method of collecting a multiphase fluid specimen, separating saidspecimen into at least two differing-density phases, and partitioningsaid phases comprising:

a. providing an open-ended container with thixotrosaid gel-like meanshaving a. specific gravity interlighter phase of a centrifugallyseparated fluid specimen within a container which comprises:

providing a container having a closed end and an open end;

initially positioning thixotropic gel-like means having a. specificgravity intermediate those of said lighter and heavier fluid phaseswithin said container in spaced relation from said open end; evacuatingand sealing said container to provide a closed fluid collection chambertherewithin; supplying a fluid specimen to said closed chamber;

pic gel-like material having a specific gravity intermediate those ofthe two phases of a fluid specimen to be collected and separated;

b. vacuum-sealing the open end of said container,

containing said gel-like material, with a needlepierceable closure;

c. drawing a specimen through said closure; d. applying centrifugalforce to said specimen and gel-like material and simultaneously forciblymoving the phases of said specimen and said gel-like material towardrelative positions within said container corresponding to theirrespective specific gravities;

. terminating said centrifugal force after said specimen -has separatedinto differing-density phases and a substantial portion of said gel-likematerial has reached a position intermediate said phases, and

. at such position, utilizing said gel-like material to partition saidseparated differing-density phases.

1. A FLUID COLLECTION AND PARTITIONING ASSEMBLY FOR COLLECTING ASPECIMEN OF BLOOD WITHIN A SEALED FLUID COLLECTION CHAMBER,CENTRIFICALLY SEPARATING THE HEAVIER AND LIGHTER FLUID PHASES OF SAIDBLOOD SPECIMEN, AND PHYSICALLY AND CHEMICALLY PARTITIONING THE SEPARATEDPHASES, COMPRISING: A. A CONTAINER HAVING AN OPEN END AND A CLOSED END;B. GEL-LIKE MEANS INITIALLY POSITIONED WITHIN SAID CONTAINER ADJACENTSAID CLOSED END FOR FORMING A TRANSVERSELY CONTINUOUS CONTACT SEAL WITHAN ANNULAR SURFACE PORTION WITHIN SAID CONTAINER AT A SUBSEQUENTLYFORMED INTERFACE BETWEEN SAID HEAVIER AND LIGHTER PHASES; C. CLOSUREMEANS FOR VACUUM-SEALING SAID OPEN END OF SAID CONTAINER AND FORDEFINING A CLOSED FLUID COLLECTION CHAMBER CONTAINING SAID GEL-LIKEMEANS THEREWITHIN, SAID CLOSURE MEANS BEING PIERCEABLE BY A NEEDLE FORSUPPLYING BLOOD TO SAID CLOSED FLUID COLLECTION CHAMBER WHICH IS ADAPTEDTO DRAW THE BLOOD SPECIMEN THEREWITHIN; D. SAID GEL-LIKE MEANS BEING ATHIXOTROPIC MATERIAL AND INCLUDING A MIXTURE OF A FLUID WHICH ISGENERALLY INERT TO BODY FLUIDS AND A POWDERED INORGANIC FILLER; AND E.SAID GEL-LIKE MEANS HAVING A SPECIFIC GRAVITY INTERMEDIATE THOSE OF SAIDLIGHTER AND HEAVIER PHASES AND BEING OF SUCH A THIXOTROPIC COMPOSITIONSUCH THAT DURING THE CENTRIFUGATION OF SAID BLOOD SPECIMEN INTO ITSCOMPONENT PHASES, SAID GEL-LIKE MATERIAL IS FLOWABLE FROM ITS INITIALPOSITION ADJACENT SAID CLOSED END TOWARD SAID SEALED OPEN END ANDEFFECTS A SEMI-RIGID SEAL AT THE INTERFACE OF SAID SEPARATED FLUIDPHASES WHICH PHYSICALLY AND CHEMICALLY PARTITIONS SAID PHASES WITHINSAID CONTAINER.
 2. A method of partitioning a heavier phase from alighter phase of a centrifugally separated fluid specimen within acontainer which comprises: a. providing a container having a closed endand an open end; b. initially positioning thixotropic gel-like meanshaving a specific gravity intermediate those of said lighter and heavierfluid phases within said container in spaced relation from said openend; c. evacuating and sealing said container to provide a closed fluidcollection chamber therewithin; d. supplying a fluid specimen to saidclosed chamber; e. subjecting said specimen and gel-like means to acentrifugal force to separate said fluid specimen into a heavier phaseand a lighter phase and simultaneously move said gel-like means towardthe interface of said phases; and f. establishing a contiNuoussemi-rigid gel-like seal across the interior of said container betweensaid heavier phase and said lighter phase within said container.
 3. Amethod of collecting a multiphase fluid specimen, separating saidspecimen into at least two differing-density phases, and partitioningsaid phases comprising: a. providing an open-ended container withthixotropic gel-like material having a specific gravity intermediatethose of the two phases of a fluid specimen to be collected andseparated; b. vacuum-sealing the open end of said container, containingsaid gel-like material, with a needle-pierceable closure; c. drawing aspecimen through said closure; d. applying centrifugal force to saidspecimen and gel-like material and simultaneously forcibly moving thephases of said specimen and said gel-like material toward relativepositions within said container corresponding to their respectivespecific gravities; e. terminating said centrifugal force after saidspecimen has separated into differing-density phases and a substantialportion of said gel-like material has reached a position intermediatesaid phases, and f. at such position, utilizing said gel-like materialto partition said separated differing-density phases.